Cargando…

Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris

Interleukin (IL)-25 (also known as IL-17E) is a distinct member of the IL-17 cytokine family which induces IL-4, IL-5, and IL-13 expression and promotes pathogenic T helper (Th)-2 cell responses in various organs. IL-25 has been shown to have crucial role between innate and adaptive immunity and als...

Descripción completa

Detalles Bibliográficos
Autores principales: Liu, Yushan, Wu, Chengsheng, Wang, Jinyu, Mo, Wei, Yu, Min
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3838785/
https://www.ncbi.nlm.nih.gov/pubmed/24100683
http://dx.doi.org/10.1007/s00253-013-5264-4
_version_ 1782478383132704768
author Liu, Yushan
Wu, Chengsheng
Wang, Jinyu
Mo, Wei
Yu, Min
author_facet Liu, Yushan
Wu, Chengsheng
Wang, Jinyu
Mo, Wei
Yu, Min
author_sort Liu, Yushan
collection PubMed
description Interleukin (IL)-25 (also known as IL-17E) is a distinct member of the IL-17 cytokine family which induces IL-4, IL-5, and IL-13 expression and promotes pathogenic T helper (Th)-2 cell responses in various organs. IL-25 has been shown to have crucial role between innate and adaptive immunity and also a key component of the protection of gastrointestinal helminthes. In this study, to produce bioactive recombinant human IL-25 (rhIL-25), the cDNA of mature IL-25 was performed codon optimization based on methylotropic yeast Pichia pastoris codon bias and cloned into the expression vector pPICZαA. The recombinant vector was transformed into P. pichia strain X-33 and selected by zeocin resistance. Benchtop fermentation and simple purification strategy were established to purify the rhIL-25 with about 17 kDa molecular mass. Functional analysis showed that purified rhIL-25 specifically bond to receptor IL-17BR and induce G-CSF production in vitro. Further annexin V-FITC/PI staining assay indicated that rhIL-25 induced apoptosis in two breast cancer cells, MDA-MB-231 and HBL-100. This study provides a new strategy for the large-scale production of bioactive IL-25 for biological and therapeutic applications.
format Online
Article
Text
id pubmed-3838785
institution National Center for Biotechnology Information
language English
publishDate 2013
publisher Springer Berlin Heidelberg
record_format MEDLINE/PubMed
spelling pubmed-38387852013-12-02 Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris Liu, Yushan Wu, Chengsheng Wang, Jinyu Mo, Wei Yu, Min Appl Microbiol Biotechnol Biotechnological Products and Process Engineering Interleukin (IL)-25 (also known as IL-17E) is a distinct member of the IL-17 cytokine family which induces IL-4, IL-5, and IL-13 expression and promotes pathogenic T helper (Th)-2 cell responses in various organs. IL-25 has been shown to have crucial role between innate and adaptive immunity and also a key component of the protection of gastrointestinal helminthes. In this study, to produce bioactive recombinant human IL-25 (rhIL-25), the cDNA of mature IL-25 was performed codon optimization based on methylotropic yeast Pichia pastoris codon bias and cloned into the expression vector pPICZαA. The recombinant vector was transformed into P. pichia strain X-33 and selected by zeocin resistance. Benchtop fermentation and simple purification strategy were established to purify the rhIL-25 with about 17 kDa molecular mass. Functional analysis showed that purified rhIL-25 specifically bond to receptor IL-17BR and induce G-CSF production in vitro. Further annexin V-FITC/PI staining assay indicated that rhIL-25 induced apoptosis in two breast cancer cells, MDA-MB-231 and HBL-100. This study provides a new strategy for the large-scale production of bioactive IL-25 for biological and therapeutic applications. Springer Berlin Heidelberg 2013-10-08 2013 /pmc/articles/PMC3838785/ /pubmed/24100683 http://dx.doi.org/10.1007/s00253-013-5264-4 Text en © The Author(s) 2013 https://creativecommons.org/licenses/by-nc/2.0/ Open Access This article is distributed under the terms of the Creative Commons Attribution License which permits any use, distribution, and reproduction in any medium, provided the original author(s) and the source are credited.
spellingShingle Biotechnological Products and Process Engineering
Liu, Yushan
Wu, Chengsheng
Wang, Jinyu
Mo, Wei
Yu, Min
Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title_full Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title_fullStr Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title_full_unstemmed Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title_short Codon optimization, expression, purification, and functional characterization of recombinant human IL-25 in Pichia pastoris
title_sort codon optimization, expression, purification, and functional characterization of recombinant human il-25 in pichia pastoris
topic Biotechnological Products and Process Engineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3838785/
https://www.ncbi.nlm.nih.gov/pubmed/24100683
http://dx.doi.org/10.1007/s00253-013-5264-4
work_keys_str_mv AT liuyushan codonoptimizationexpressionpurificationandfunctionalcharacterizationofrecombinanthumanil25inpichiapastoris
AT wuchengsheng codonoptimizationexpressionpurificationandfunctionalcharacterizationofrecombinanthumanil25inpichiapastoris
AT wangjinyu codonoptimizationexpressionpurificationandfunctionalcharacterizationofrecombinanthumanil25inpichiapastoris
AT mowei codonoptimizationexpressionpurificationandfunctionalcharacterizationofrecombinanthumanil25inpichiapastoris
AT yumin codonoptimizationexpressionpurificationandfunctionalcharacterizationofrecombinanthumanil25inpichiapastoris